Innate immune response to viral infection Cytokine - sepeap

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Cytokine
journal homepage: www.elsevier.com/locate/issn/10434666
Review Article
Innate immune response to viral infection
Shohei Koyama a,b , Ken J. Ishii a,c , Cevayir Coban a,b , Shizuo Akira a,b, *
a Laboratory of Host Defense, WPI Immunology Frontier Research Center, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan
b Department of Host Defense, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan
c Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan
article
info
abstract
Article history:
Received 2 June 2008
Accepted 9 June 2008
Available online xxxx
Keywords:
Pattern recognition receptors
Type I interferons
Nucleic acids
In viral infections the host innate immune system is meant to act as a first line defense to prevent viral
invasion or replication before more specific protection by the adaptive immune system is generated. In
the innate immune response, pattern recognition receptors (PRRs) are engaged to detect specific viral
components such as viral RNA or DNA or viral intermediate products and to induce type I interferons
(IFNs) and other pro-inflammatory cytokines in the infected cells and other immune cells. Recently these
innate immune receptors and their unique downstream pathways have been identified. Here, we summarize
their roles in the innate immune response to virus infection, discrimination between self and viral
nucleic acids and inhibition by virulent factors and provide some recent advances in the coordination
between innate and adaptive immune activation.
Ó 2008 Published by Elsevier Ltd.
1. Introduction
* Corresponding author. Address: Laboratory of Host Defense, WPI Immunology
Frontier Research Center, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-
0871, Japan. Fax: +81 6 6879 8305.
E-mail address: sakira@biken.osaka-u.ac.jp (S. Akira).
All living organisms have developed several kinds of mechanisms
to protect themselves from invasion by exogenous microorganisms,
including viruses. Although the production of neutralizing antibodies
and activation of cytotoxic T lymphocytes (CTL) or natural killer
(NK) cells are essential for a specific and effective antiviral immune
response, other host cells also possess some immune mechanism to
prevent viral infection.
Although multiple cytokines and chemokines are produced by
several kinds of host cells in viral infection, type I IFNs are the principal
cytokines involved in the antiviral response. Type I IFNs
include multiple IFN-a isoforms, a single IFN-b, and other members,
such as IFN-e, -j, -x and so on [1]. In contrast to type II IFN
(IFN-c), which is exclusively produced by T cells and NK cells, type
I IFNs can be produced by all nucleated cells in response to virus
infection. Type III IFNs, comprised of IFN-k1, k2 and k3, have also
recently been identified [2]. These IFNs each have different receptors
but share downstream signaling molecules and regulate the
same genes. IFNs have pleiotropic functions. They increase the
expression of intrinsic proteins including TRIM5a, Fv, Mx, eIF2a
and 2 0 –5 0 OAS, and induce apoptosis of virus-infected cells and cellular
resistance to viral infection [3]. In addition they activate NK
cells and dendritic cells (DC) and induce the activation of the adaptive
immune system [4]. The expression of type I IFN and cytokine
genes is regulated by an intracellular signaling pathway that is
activated by germline-encoded PRRs. These receptors recognize
molecular patterns specific to microorganisms, such as viral genome
nucleic acids. Nucleic acids such as DNA and RNA are essential
components of all living organisms, so discrimination between self
and non-self nucleic acids is essential especially in virus infection.
Recent advances in research into innate immunity have revealed
that this discrimination relies, to a great extent, on PRRs including
Toll-like receptors (TLRs), retinoic acid-inducible gene I (RIG-I)-like
receptors (RLRs), and nucleotide-binding oligomerization domain
(NOD)-like receptors (NLRs). Here, we review the current understanding
of innate immune recognition of viruses and discrimination
between self and viral nucleic acids, and provide some
recent advances in coordination between innate immune signaling
and adaptive immune activation.
2. Innate immune receptors for virus sensing
2.1. Endosomal TLRs in DCs
Several kinds of viruses utilize host endocytic pathways at the
cell entry phase or budding, so they are inevitably surveyed by
endosomal innate immune sensors. Endosomal TLRs, including
TLR3, TLR7, TLR8 and TLR9, share the property of being activated
by nucleic acids. Their expression can be increased by type I IFNs
but their distribution is restricted. TLR7 and TLR9 are highly
expressed in plasmacytoid DCs (pDCs) which are expert cells
known to produce a large amount of type I IFNs in response to
1043-4666/$ - see front matter Ó 2008 Published by Elsevier Ltd.
doi:10.1016/j.cyto.2008.07.009
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virus infection. TLR3 is expressed more widely, but is mainly expressed
on conventional DCs (cDCs) [5]. The function of TLR8 is
not clearly known yet.
Recently endoplasmic reticulum (ER) protein UNC93B1 turned
out to be essential for trafficking of TLR7 and TLR9 from ER to
endosome [6], but what triggers this TLR-trafficking from ER to
endosome before viral recognition by TLRs remains to be elucidated.
A recent report suggests that TLR-sorting to the ligand
may utilize autophagy, which is a cellular process for recycling
cytosolic compartments and, in the case of pDC, eliminating exogenous
pathogen. Although cytoplasmic vesicular somatitis viruses
(VSV) in pDC are thought to be trapped by autophagosome
expressing ATG5 and detected by TLR7 in lysosomes for a type I
IFN response [7], in non-immune cells ATG5 suppresses the type
I IFN response by interaction with caspase recruitment domains
(CARDs) presented by RIG-I and IFN-b promoter stimulator-1
(IPS-1) [8] (Fig. 1).
2.2. Endosomal recognition of viral RNA by TLRs
TLR7 recognizes several kinds of RNA viruses, including
orthomyxoviruses in pDCs. TLR7 signals through a TIR domaincontaining
adapter, myeloid differentiation factor 88 (MyD88).
Upon exposure to its ligand, MyD88 forms a complex with interleukin-1-receptor
(IL-1R)-associated kinase-4 (IRAK-4), IRAK-1, tumor
necrosis factor-receptor associated factor 3 (TRAF3), TRAF6,
Ikka and IRF-7 [9–11]. Following the formation of this signaling
complex, IRF7 and nuclear factor-kappa B (NF-jB) are activated,
which results in the production of type I IFNs and cytokines (Fig. 1).
TLR3 recognizes double-stranded (ds)RNA and triggers a signaling
pathway via a TIR domain-containing adapter inducing IFN-b
(TRIF) (also known as TICAM-1) [12,13]. TRIF associates with
TRAF3 and TRAF6 via TRAF-binding motifs which exist in its N-terminal
portion and also with receptor interacting protein (RIP) 1
and RIP3 via RIP homotypic interaction motif (RHIM) [14,15].
TRAF6 and RIP1 activate NF-jB while TRAF3 activates TRAF family
member-associated NK-jB activator (TANK)-binding kinase 1
(TBK1) and inducible IjB kinase (IKK-i). Activation of these pathways
triggers antiviral responses (Fig. 1). TLR3 also activates the
phosphatidylinositol-3 kinase (PI3K) pathway [16]. Tyrosine phosphorylation
of TLR3 induces PI3K recruitment to the receptor and
subsequent activation of Akt leads to activation of IRF-3. TLR3
plays an important role in the pathogenesis of RNA virus infections
in vivo. For example, TLR3 deficient mice are resistant to infection
Fig. 1. RNA sensing in virus infection. TLR3 recognizes dsRNA and triggers a signaling pathway via a TRIF. TRIF associates with TRAF3, TRAF6 and RIP1. TRAF6 and RIP1
activate NF-kB and AP-1 while TRAF3 activates TBK1/IKK-i and is followed by a type I IFN response. Both RIG-I and MDA5 associate with an adapter protein IPS-1. IPS-1
localizes on the outer mitochondrial membrane and the CARD of it interacts with that of RIG-I or MDA5. IPS-1 associates with TRAF3 which induces the production of type I
IFNs and FADD which induces activation of NF-kB. TLR7 signals through MyD88. MyD88 forms a complex with IRAK-4, IRAK-1, TRAF3, TRAF6, Ikka and IRF-7 and this complex
is recruited to the TLR by ligand stimulation. Downstream of this signaling complex, IRF7 and NF-kB are activated and this is followed by the production of type I IFNs and
cytokines. ER protein UNC93B1 plays a key role in trafficking of TLR7 from ER to endosome, but the triggers are unknown. In the case of VSV recognition in pDC, the virus is
trapped by autophagosome expressing ATG5 and detected by TLR7 in the lysosome for production of a type I IFN response. Moreover in non-immune cells ATG5 suppresses
the type I IFN response via RIG-I or IPS-1 inhibition. Yellow rectangles (TIR, CARD) indicate protein–protein interaction regions for downstream signaling.
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with West Nile virus [17] and influenza virus [18]. In both cases
inflammatory responses are decreased in TLR3 deficient mice,
which suggests that an excess production of cytokines is rather
harmful for the survival of mice.
2.3. Intracellular recognition of viral RNA by RLRs
Recently, three homologous DExD/H box RNA helicases, RIG-I,
melanoma differentiation-associated gene 5 (MDA5) and LGP2,
were identified as cytoplasmic sensors of virus RNA, named here
as RIG-like receptors (RLRs). RIG-I and MDA5 play a major role in
recognition of RNA viruses in cDCs, macrophages and fibroblasts.
RIG-I and MDA5 share two N-terminal CARDs followed by an
RNA helicase domain [19] while LGP2 lacks a CARD. RIG-I binds
5 0 -triphosphorylated single-stranded (ss)RNA [20] and short
dsRNA [21] and stimulates production of type I IFNs. By contrast,
MDA5 preferentially recognizes longer-dsRNA, including synthetic
poly-IC [22]. RIG-I recognizes a variety of RNA viruses
including influenza virus, VSV and Japanese encephalitis virus
(JEV) while MDA5 recognizes picorna family such as encephalomyocarditis
virus (EMCV), Theiler’s virus and Mengo virus [22].
Therefore, RIG-I and MDA5 deficient mice are highly susceptible
to VSV and EMCV, respectively. In addition it was indicated that
the antiviral protein RNase L, which can cleave and turn a singlestranded
portion of not only viral but also self RNA into preferentially
double-stranded form RNA, is a ligand for RIG-I and
MDA5 [23].
The CARDs of RIG-I and MDA5 are responsible for initiating the
signaling pathway. Both RIG-I and MDA5 associate with an adapter
protein IPS-1 also known as MAVS, VISA or CARDIF, which also
contains an N-terminal CARD [24–27]. IPS-1 localizes on the outer
mitochondrial membrane. The IPS-1 CARD interacts with that of
RIG-I or MDA5. IPS-1 then associates with TRAF3, followed by activation
of TBK1 and Ikk-i. These kinases phosphorylate IRF3 and
IRF7 and induce type I IFN production [28,29]. IPS-1 also interacts
with Fas-associated death domain-containing protein (FADD) and
leads to activation of NF-jB through cleavage of caspase-8/-10
[30] (Fig. 1).
2.4. Differential role of TLR and RLR in antiviral responses
Occasionally both TLR and RLR are engaged for sensing the same
dsRNA or ssRNA. Normally dsRNA does not exist in host cells but in
virus infection it is detected as not only a viral structure but also as
a byproduct of viral replication. dsRNA activates macrophages and
dendritic cells via TLR3 to secrete pro-inflammatory cytokines,
especially IL-12. However, type-I IFNs are produced by virus-infected
cells such as fibroblasts by TLR3 independently. MDA5 recognizes
synthetic dsRNA, poly-IC, and the ssRNA virus, EMCV,
which generates dsRNA during replication, and induces the type-I
IFN response [22,31]. Poly-IC is neither capable of inducing an innate
immune response nor of working as an adjuvant in TRIF/
IPS-1 double knockout mice [32].
In contrast to dsRNA, ssRNA abundantly exists not only in
pathogens but also in host cells. ssRNA is recognized by TLR7
(or TLR8 in humans) and RIG-I in a cell-type specific manner.
Although TLR7 and TLR8 recognize GU of AU rich sequences of
ssRNA viruses such as influenza virus and HIV, through TLR7
expressing cells such as pDC, or TLR8 expressing cells such as
myeloid DC or monocytes, it is unclear whether its sequence
specificity is dependent on the receptor or the cell [33]. In contrast
to TLR7, RIG-I is expressed in most cell types. As described
previously, RIG-I recognizes 5 0 -triphosphorylated ssRNA [20]. In
the case of influenza A virus infection, its negative-sense ssRNA
genome is recognized by TLR7 expressed in pDCs and a signal is
transmitted through its adaptor protein MyD88 [34]. On the other
hand, it is recognized by RIG-I ubiquitously expressed in most cell
types, such as fibroblasts or cDCs in vitro [35], and probably by
the alveolar macrophage in vivo [36], via its adaptor protein
IPS-1. In mouse lungs after intranasal infection of influenza virus
both TLR7/MyD88 and RIG-I/IPS-1 pathways concurrently control
the type-I IFN response [37].
2.5. Endosomal and intracellular recognition of viral DNA
TLR9 recognizes unmethylated DNA with a CpG motif (CpG-
DNA) and DNA viruses, including herpes simplex virus (HSV)-1,
HSV-2 and cytomegalovirus (CMV) in pDCs. TLR9 shares the adapter
protein MyD88 and the downstream signaling pathway with
TLR7. cDCs and macrophages also respond to CpG-DNA and produce
small amounts of IFN-b through IRF-1 activation rather than
IRF-3 or IRF-7 activation [38].
Recently, it was reported that genomic DNA of viruses, such as
adenovirus, vaccinia virus and HSV [39–41], could be also recognized
in a TLR9-independent manner, using an as yet unknown
recognition mechanism in the cytoplasm [42,43]. In this case,
DNA which has entered the cytoplasm activates the infected cells
via TBK1 and IRF3 [44]. Actually the source of DNA is not restricted
to viruses; but it can also come from bacteria and damaged host
cells. The activity of this DNA is more potent in ds right-hand B-
form DNA than in left-handed Z-form DNA, while ssDNA displays
no activity [40,44,45] (Fig. 2). In addition DAI (also known as
ZBP1 or DLM1) which contains two Z-DNA binding domains, was
shown to be a potential cytoplasmic DNA sensor [46]. However,
DAI KO mice induced a normal type I IFN response in vitro and
in vivo after B-DNA stimulation and also indicated DNA-vaccine-induced
adaptive immune responses, suggesting its role is redundant
[47]. Potential cytoplasmic DNA sensors still remain to be
elucidated.
2.6. The recognition of viral components at the cell surface
In addition to the endosomal TLRs, TLR2 and TLR4 have also
been suggested to be involved in recognition of viruses. TLR2 has
been shown to detect components of measles virus, HSV and hepatitis
C virus (HCV) [48–50], while TLR4 produces a response to retrovirus
and respiratory syncytial virus (RSV) [51,52]. While viral
proteins recognized by these surface TLRs trigger pro-inflammatory
responses, their contribution to either protective or pathological
immune responses largely depends on the type of virus, route
of infection, and other host factors [53].
2.7. NLRs mediates innate immune activation by intracellular viral
nucleic acids
NLR proteins are comprised three motifs, C-terminal LRRs, central
nucleotide-binding domain and N-terminal signaling domaincontaining
CARDs, and Pyrin domain or baculovirus IAP repeats
[54]. Cryopyrin/NALP3 was shown to recognize both ssRNA and
dsRNA of viral origin or their synthetic versions and to induce caspase-1
activation via apoptosis-associated speck-like protein containing
a caspase-activating and recruitment domain (ASC)
[55,56]. In addition, some NLRs participate in nucleic acid-mediated
innate immune activation through caspase-1 activation
[56,57], and NF-jB activation towards IFN-I production via a synergistic
pathway activated by NOD2 [58].
3. Discrimination between self and viral nucleic acids
The innate immune systems for virus sensing described above
miraculously detect the invasion of pathogens such as viruses,
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Fig. 2. DNA sensing in virus infection. TLR9 recognizes CpG-DNA and DNA viruses, including HSV-1, HSV-2 and CMV. TLR9 shares MyD88 and a downstream signaling
pathway with TLR7 and ER protein UNC93B1 also plays a key role in trafficking of TLR9 from ER to endosome. In TLR9-independent DNA sensing, DNAs enter into the
cytoplasm which activates the infected cells via TBK1 and IRF3 but the receptor and adaptor involved are still unknown. Their activities are more potent in ds right-hand B-
form DNA than in left-handed Z-form DNA.
but are silent in normal conditions. As dsRNA generated by viral
replication and virus DNA rich in CpG motifs are not normally
found in our body, it is easy to consider that our innate immune
system recognizes them as foreign molecules. In contrast to these
nucleic acids, ssRNA abundantly exists not only in pathogens but
also in host cells. It is, therefore, essential for the host to detect
and discriminate viral ssRNA from self ssRNA. However, the mechanism
of this robust discrimination is not fully understood. For
example, although the 5 0 triphosphate on many ssRNAs of viruses
is absent from mRNA and transfer RNA but is found in ribosomal
RNA, which abundantly exists in host cells, only the 5 0 triphosphate
on viruses can induce RIG-I activation [59,60]. Recently RNase L
activation in infected host cells was shown to generate small self
RNAs which can induce an innate immune response via RLRs
[61]. In addition, it is known that TLR7/8 and TLR9 also recognize
host RNA and DNA. Therefore, it is necessary for absolute discrimination
by the host innate immune system to recognize some additional
factors such as the methylation state, certain sequences and
intracellular localization of RNA, or restricted endosomal expression
of TLRs for viral recognition where host nucleic acids have limited
access [62–64].
4. Virulent factor for inhibition of host immune response
Viruses have developed several kinds of immune evasion strategies
to proliferate within host cells. Their main target is the type I
IFN response. Viruses can inhibit type I IFNs by many strategies i.e.
inhibition of IFN synthesis, interference of IFN receptor signaling
and so on [65]. For example, vaccinia virus E3L and influenza virus
NS1 which possess a dsRNA-binding site are thought to inhibit
type I IFN production through dsRNA sequestration [66,67]. As
E3L also possesses a DNA-binding site, it might sequester viral
DNA from host DNA sensing [68]. NS1 protein was also indicated
to inhibit the function of IPS1 and RIG-I [69]. Viruses without such
abilities to suppress the host type I IFN responses are generally low
pathogenic and available for vaccine strains.
5. Adaptive immunity against viruses through innate immune
signaling pathway
Recent advances in the understanding of innate immunity show
that the activation of the innate immune system is essential for
subsequent adaptive immune responses including specific antibody
production and CTL activation which play a key role in protection
against virus infection. A recent report indicated that the
adaptive immune response elicited by inactivated whole influenza
virus vaccine containing viral ssRNA was strictly governed by the
TLR7/MyD88 pathway, but not by the RIG-I/IPS-1 pathway,
although both pathways concurrently controlled the innate immune
response [37]. However, the innate immune response elicited
by a DNA vaccine containing CpG-DNA was dependent on
TBK1, but not on the TLR9/MyD88 pathway [47]. These results sug-
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gested that the innate immune pathway engaged for protective
immunity against virus infection was different according to the
source of antigen.
6. Conclusion
This review has illustrated the recent progress in understanding
how a host discriminates between viruses and self components by
innate immune receptors and elicits an inflammatory response
involving type I IFNs and other cytokines. Much remains to be clarified
about the complex interplay between host and virus, but elucidating
such mechanisms in detail is essential for not only the
development of a clinical approach such as nucleic acid-based
immunotherapy and TLR based vaccine adjuvant but also the
understanding of the pathogenesis of diverse viral diseases.
Acknowledgments
We thank members of Akira Laboratory, Prof. Toshihiro Horii
and his laboratory members for discussions and contributions to
the work discussed here. This work was supported by grants from
the Ministry of Education, Culture, Sports, Science and Technology
in Japan.
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